Neurons are highly polarized cells with distinct domains that mediate specialized functions. Incoming signals from other neurons are generally detected at synapses localized to the dendrites and cell body. The outgoing signal is conducted via the axon to the nerve terminal, where the resulting depolarization causes an influx of calcium and the release of neurotransmitter by fusion of synaptic vesicles with the presynaptic plasma membrane. These functional differences are reflected in differences in the distributions of specific proteins and organelles. For example, neurotransmitter receptors must be targeted to postsynaptic membrane specializations at dendrites, while synaptic vesicles and their constituent proteins are concentrated at the nerve terminal. The specific aims of this proposal are designed to determine the mechanism mediating the specific targeting of synaptic vesicle proteins to the nerve terminal and, ultimately, their assembly into synaptic vesicles. Hippocampal neurons grown at low density in tissue culture develop the characteristic polarized morphology of neurons in vivo. Synaptic vesicle proteins are enriched in the axon, while proteins such as the transferrin receptor are localized to the somatodendritic region. We will use this system to investigate the amino acid sequences that are responsible for the targeting of synaptic vesicles to the axon, and ultimately to synaptic vesicles. We will identify the signals responsible for the sorting of the synaptic vesicle protein synaptobrevin to the nerve terminal. We will determine if sorting to the axonal or somatodendritic domain is mediated by different determinants, and how those determinants interact to create a polarized distribution of proteins in the neuron. Finally, we will determine whether sorting determinants are conserved between different types of regulated secretory vesicles. With these results in hand, we will have taken the first step towards understanding the complex mechanism that results in the formation of synaptic vesicles during development and during recycling at the nerve terminal.